Thyroid hormones function in diverse and important physiological roles throughout the life an organism, ranging from the development of multiple organ systems to the regulation of metabolic rate. The thyroid is a unique collection of epithelial follicles that are organized to secrete the scaffold for hormone production, thyroglobulin, as well as iodinate and store the resultant thyroid hormone precursor. They also respond to the signaled need for thyroid hormone by processing and releasing thyroid hormones. Processes relevant to proper hormone synthesis take place within the follicle and also are compromised in many disease states. Our long-term goal is to understand how the thyroid follicular lumen composition is maintained. Our group recently positively established the presence of Cystic Fibrosis Transmembrane Conductance Regulator (CFTR) in thyroid, where it can mediate thyroid anion secretion via an apically localized, cAMP-stimulated, conductive pathway for anion exit. Considering the importance of thyroid hormone to so many essential physiological processes, it is critical to identify and understand the role of CFTR in thyroid function. The present proposal aims to understand and develop a model describing the regulatory interaction between CFTR and SLC5A8, a solute transporter that is highly expressed at the apical membrane of the thyroid follicular epithelial cell. Our studies indicate that SLC5A8 activity depends on activation of CFTR, thus suggesting functional interactions. Among the reported functions for SLC5A8 are roles in iodide transport, short-chain fatty acid transport and tumor suppression. All are of potentially high relevance to thyroid health and disease and yet surprisingly little is known about them. CFTR regulation of other solute transport molecules of a different gene family (SLC26) is well-studied. Current models for CFTR regulation of SLC26s indicate protein-protein interactions that are facilitated by scaffolding molecules. SLC5A8 contains a well-studied protein interaction motif that is also found in CFTR, yet no information about its functional role exists in the literature. We hypothesize that CFTR regulates SLC5A8 via protein-protein interactions. Preliminary data using heterologous co-expression of CFTR and SLC5A8 in amphibian and mammalian cell model systems align with the predictions of the hypothesis. The proposed research strategy combines well-established methods to define both the molecular requirements for CFTR and SLC5A8 interaction(s) within macromolecular complex (es), as well as their regulatory crosstalk, i.e. the functional outcomes of interaction. Our proposed studies utilize complementary biochemical/cell biological approaches and functional electrophysiological and fluorimetric measurements. The resulting information will be integrated into models that can be iteratively challenged, developed and refined. The present proposal will be the first to test the hypothesis that both CFTR and SLC5A8 associate within a common macromolecular complex. Not only will all emergent findings be new and novel, but they potentially will impact the direction of multiple fields.